Recent studies indicate that the diaphragm becomes profoundly weak in critically ill patients. Diaphragm weakness, in turn, increases the duration of mechanical ventilation, increases patient mortality, and is a major contributor to the high cost of taking care of critically ill, mechanically ventilated patients in the VA health care system. Our own work indicates that infections are a major cause of the development of severe diaphragm weakness in critically ill patients. It is also known that infections elicit increases in diaphragm free radical generation, and that heightened free radical generation contributes to the development of weakness. The precise mechanism(s) by which infections activate free radical generation in skeletal muscle are not known, however. Moreover, currently there is no pharmacological treatment to prevent or reverse infection induced diaphragm weakness in critically ill patients. The purpose of the present project is to discover the mechanisms by which infections increase diaphragm free generation and to employ this mechanistic information to define novel translatable pharmacological treatments that can be used to prevent and reverse diaphragm weakness in critically patients. Our central hypothesis is that infection induced diaphragm dysfunction is primarily a consequence of the sequential activation of neutral sphingomyelinase 2, cytosolic phospholipase A2 (cPLA2), mitochondrial free radical generation, and proteolytic pathways (e.g. calpain). The planned studies will test this hypothesis and use this information to define new therapies that can be quickly translated into clinical usage. Aim 1 experiments will delineate the specific mechanism(s) by which infections induce heightened free radical generation in skeletal muscle. Experiment 1.1 will test the hypothesis that infections first activate neutral sphingomyelinase 2 in skeletal muscle. Experiment 1.2 will test the hypothesis that infection induced activation of neutral sphingomyelinase 2 subsequently activates skeletal muscle cPLA2, which, in turn, induces an increase in mitochondrial free radical production. Aim 2 experiments will determine the specific skeletal muscle proteolytic pathways activated (Experiment 2.1) and cellular proteins degraded (Experiment 2.2) as a consequence of infection induced activation of cPLA2 and mitochondrial free radical generation. Aim 3 experiments will determine if novel pharmacological agents which block cPLA2 activation and mitochondrial free radical generation prevent loss of diaphragm function in the cecal ligation perforation animal model of sepsis. We will study an agent which blocks activation of cPLA2 (taurine) in Experiment 3.1, a direct cPLA2 inhibitor (CDIBA) in Experiment 3.2, and an inhibitor of mitochondrial free radical generation (SS31) in Experiment 3.3.